Piperazinyl-or piperidinylamine-sulfamic acid amides as inhibitors of steroid sulfatase

ABSTRACT

Piperazinyl- or piperidinylamine-sulfamic acid amides and their use for the manufacture of a medicament in diseases mediated by the action of steriod sulfatase.

The present invention relates to sulfamic acid amides, e.g. useful inthe treatment of disorders mediated by the action of steroid sulfatase.

In one aspect the present invention provides a compound of formula

wherein either

-   -   R₁ and R₂ together with the nitrogen atom to which they are        attached are piperazinyl, wherein the second nitrogen atom is        substituted by (C₁₋₆)alkoxycarbonyl or by (C₆₋₁₈)aryl, which        (C₆₋₁₈)aryl is substituted by one or more halogen,        (C₁₋₆)haloalkyl, e.g. CF₃, aminocarbonyl; or    -   R₁ is hydrogen and R₂ is piperidinyl, attached via a carbon atom        of the piperidinyl ring, wherein the nitrogen atom is        substituted by (C₁₋₆)alkoxycarbonyl or by (C₆₋₁₈)aryl, and    -   R₃ is (C₆₋₁₈)aryl or (C₆₋₁₈)aryl(C₁₋₄)alkyl, which aryl is        substituted by one or more halogen, aminocarbonyl, or        (C₁₋₆)haloalkyl.

If not otherwise defined herein (C₆₋₁₈)aryl, e.g. phenyl, such as(C₆₋₁₈)aryl substituted by one or more halogen, (C₁₋₆)haloalkyl, e.g.CF₃, or aminocarbonyl. (C₁₋₆)Alkoxycarbonyl includes unsubstituted orsubstituted (C₁₋₆)alkoxycarbonyl, e.g. alkoxycarbonyl wherein the alkylgroup is substituted by (C₆₋₁₈)aryl, such as phenyl, e.g.benzyloxycarbonyl. Halogen includes fluoro, chloro, bromo, iodo, e.g.fluoro, chloro. (C₆₋₁₈)Aryl(C₁₋₄)alkyl is preferably phenyl(C₁₋₄)alkyl,e.g. phenylethyl, wherein phenyl is substituted as described forsubstituted (C₆₋₁₈)aryl.

Preferably in a compound of formula I

either

-   -   R₁ and R₂ together with the nitrogen atom are piperazinyl,        wherein the second nitrogen atom is substituted by        -   (C₁₋₆)alkoxycarbonyl, e.g. tert.butoxycarbonyl,            benzyloxycarbonyl,        -   phenyl substituted by aminocarbonyl, (C₁₋₆.)haloalkyl, or    -   R₁ is hydrogen and R₂ is piperidinyl, attached via a carbon atom        of the piperidinyl ring, wherein the nitrogen atom is        substituted by        -   (C₁₋₆)alkoxycarbonyl, e.g. tert.butoxycarbonyl,            benzyloxycarbonyl,        -   phenyl substituted by aminocarbonyl, (C₁₋₆)haloalkyl, and    -   R₃ is    -   phenyl substituted by one or more        -   halogen,        -   (C₁₋₆)haloalkyl, e.g. CF₃,        -   aminocarbonyl, or    -   (C₁₋₄)alkyl, substituted by phenyl, which phenyl is substituted        by one or more        -   halogen,        -   (C₁₋₆)haloalkyl, e.g. CF₃,        -   aminocarbonyl.

In another aspect the present invention provides a compound of formulaI, which is selected from the group consisting of the compounds offormula

wherein

-   a. R_(3ss) is 3,5-bis(trifluoromethyl)phenyl and R_(4ss) is    2-aminocarbonyl-5-trifluoromethylphenyl,-   b. R_(3ss) is 2,3-dichlorophenyl, R_(4ss) is    2-aminocarbonyl-5-trifluoromethylphenyl,-   c. R_(3ss) is 3,5-dichlorophenyl R_(4ss) is    2-aminocarbonyl-5-trifluoromethylphenyl, and-   d. R_(3ss) is 3,5-bis(trifluoromethyl)phenyl and R_(4ss) is    tert.butoxycarbonyl;-   preferably R_(3ss) is 3,5-bis(trifluoromethyl)phenyl.

In another aspect the present invention provides a compound of formulaI, which is selected from the group consisting of compounds of formula

wherein

-   a. R_(3s) is 3,5-bis(trifluoromethyl)phenyl and R_(4s) is    tert.butoxycarbonyl,-   b. R_(3s) is 2,3-dichlorophenyl and R_(4s) is tert.butoxycarbonyl,-   c. R_(3s) is 3,5-dichlorophenyl and R_(4s) is tert.butoxycarbonyl,-   d. R_(3s) is 3,5-bis(trifluoromethyl)phenyl and R_(4s) is    benzyloxycarbonyl,-   e. R_(3s) is 2,3-dichlorophenyl and R_(4s) is benzyloxycarbonyl,-   f. R_(3s) is 3,5-dichlorophenyl and R_(4s) is benzyloxycarbonyl,-   g. R_(3s) is 3,5-dichlorophenyl and R_(4s) is benzyloxycarbonyl,-   h. R_(3s) is 3,5-bis(trifluoromethyl)phenyl and R_(4s) is    2-aminocarbonyl-5-trifluoromethylphenyl,    -   i. R_(3s) is 3,5-dichlorophenyl and R_(4s) is is        2-aminocarbonyl-5-trifluoromethylphenyl,-   j. R_(3s) is 2,3-dichlorophenyl and R_(4s) is is    2-aminocarbonyl-5-trifluoromethylphenyl, and    -   k. R_(3s) is 2-(3,5-bis(trifluoromethyl)phenyl)ethyl and R_(4s)        is tert.butoxycarbonyl.

Compounds provided by the present invention are hereinafter designatedas “compound(s) of (according to) the present invention”. A compound offormula I includes a compound of formula I_(s) and a compound of formulaI_(ss). Each single substituent defined above in a compound of thepresent invention may be per se a preferred substituent, independentlyof the other substituents defined. A compound of the present inventionincludes a compound in any form, e.g. in free form, in the form of asalt, in the form of a solvate and in the form of a salt and a solvate.

In another aspect the present invention provides a compound of thepresent invention in the form of a salt.

Such salts include preferably pharmaceutically acceptable salts,although pharmaceutically unacceptable salts are included, e.g. forpreparation/isolation/purification purposes.

A salt of a compound of the present invention includes a metal salt, anacid addition salt or an amine salt. Metal salts include for examplealkali or earth alkali salts; acid addition salts include salts of acompound of the present invention with an acid, e.g. HCl; amine saltsinclude salts of a compound of the present invention with an amine. Acompound of the present invention in the form of a salt is preferably ametal salt.

A compound of the present invention in free form may be converted into acorresponding compound in the form of a salt; and vice versa. A compoundof the present invention in free form or in the form of a salt and inthe form of a solvate may be converted into a corresponding compound infree form or in the form of a salt in non-solvated form; and vice versa.

A compound of the present invention may exist in the form of isomers andmixtures thereof; e.g. optical isomers, diastereoisomers, cis/transconformers. A compound of the present invention may e.g. containasymmetric carbon atoms and may thus exist in the form of enatiomers ordiastereoisomers and mixtures thereof, e.g. racemates. Any asymmetriccarbon atom may be present in the (R)-, (S)- or (R,S)-configuration,preferably in the (R)- or (S)-configuration.

Isomeric mixtures may be separated as appropriate, e.g. according, e.g.analogously, to a method as conventional, to obtain pure isomers. Thepresent invention includes a compound of the present invention in anyisomeric form and in any isomeric mixture.

The present invention also includes tautomers of a compound of formulaI, where tautomers can exist.

Any compound described herein, e.g. a compound of the present invention,may be prepared as appropriate, e.g. according, e.g. analogously, to amethod as conventional, e.g. or as specified herein.

In another aspect the present invention provides a process for theproduction of a compound of the present invention comprising the stepsof

either

-   A. reacting a compound of formula    -   wherein R₁ and R₂ are as defined above,    -   with a compound of formula    -   wherein R₃ is as defined above, e.g. in an activated form, such        as In the form of an carboxylic acid chloride, e.g. or reacting        in the presence of a coupling agent; or-   B1. reacting a compound of formula II wherein R₁ and R₂ together    with the nitrogen atom to which they are attached are unsubstituted    piperazinyl with a compound of formula III, wherein R₃ is as defined    above, e.g. in an activated form, such as in the form of an    carboxylic acid chloride, e.g. or reacting in the presence of a    coupling agent, to obtain a compound of formula    -   wherein R₃ is as defined above; and-   B2. reacting a compound of formula IV with an optionally substituted    (C₆₋₁₈)fluoroaryl, e.g. a fluorophenyl, wherein optional    aryl-substitutents are as defined in a compound of formula I, in the    presence of a base, e.g. K₂CO₃, to obtain a compound of formula I,    wherein R₁ and R₂ together with the nitrogen atom to which they are    attached are piperazinyl which is substituted at the second nitrogen    atom by optionally substituted (C₆₋₁₈)aryl; or-   C1. reacting a compound of formula    -   wherein R_(SUB) is (C₁₋₆)alkoxycarbonyl, with benzaldehyde in        the presence of NaBH₄, to obtain a compound of formula    -   wherein R_(SUB) is as defined above and Bnz is benzyl,-   C2. reacting a compound of formula VI with H₂N—SO₂—NH₂ to obtain a    compound of formula    -   wherein R_(SUB) and Bnz are as defined above,-   C3. reacting a compound of formula VII with R₃—COOH, wherein R₃ is    as defined above, e.g. wherein the carboxyl group is in an activated    form, to obtain a compound of formula    -   wherein R_(SUB), Bnz and R₃ are as defined above,-   C4. reacting a compound of formula VIII with etheric HCl to obtain a    compound of formula    -   wherein Bnz and R₃ are as defined above,-   C5. reacting a compound of formula IX with an optionally substituted    (C₆₋₁₈)fluoroaryl, e.g. a fluorophenyl, wherein optional    aryl-substitutents are as defined in a compound of formula I, in the    presence of a base, e.g. K₂CO₃, to obtain a compound of formula X,    which is a compound of formula IX, wherein Bnz and R₃ are as defined    above and the nitrogen of the piperidinyl is substituted by    optionally substituted (C₆₋₁₈)aryl,-   C6. hydrogenating a compound obtained in step C5, in the presence of    a palladium catalyst, to obtain a compound of formula I, wherein R₁    is H, R₂ is piperidine substituted by optionally substituted    (C₆₋₁₈)aryl, and R₃ is as defined above,    and isolating a compound of formula I obtained in reaction A,    reaction step B2, or reaction step C6 from the reaction mixture.

Compounds of formula II, III, IV, V, VI, VII, VIII, IX and X areintermediates (starting materials) in the production of a compound offormula I. In such intermediates (starting materials) functional groups,if present, optionally may be in protected form or in the form of asalt, if a salt-forming group is present. Protecting groups, optionallypresent, may be removed at an appropriate stage, e.g. according, e.g.analogously, to a method as conventional. A compound of formula I thusobtained may be converted into another compound of formula I, e.g. or acompound of formula I obtained in free form may be converted into a saltof a compound of formula I and vice versa.

The above reactions may be carried out as appropriate, e.g. inappropriate solvent and at appropriate temperatures, e.g. according,e.g. analogously, to a method as conventional or according to a methodas described herein. Intermediates (starting materials) are known or maybe obtained appropriately, e.g. according to a method as conventional,e.g. or as described above. Any compound described herein, e.g. acompound of the present invention, may be prepared as appropriate, e.g.according, e.g. analogously, to a method as conventional, e.g. or asdescribed herein.

Steroidal hormones in particular tissues are associated with severaldiseases, such as tumors of breast, endometrium and prostate anddisorders of the pilosebaceous unit, e.g. acne, androgenetic alopecia,and hirsutism. Important precursors for the local production of thesesteroid hormones are steroid 3-O-sulfates which are desulfated by theenzyme steroid sulfatase in the target tissues. Inhibition of thisenzyme results in reduced local levels of the corresponding activesteroidal hormones, which is expected to be of therapeutic relevance.Furthermore, steroid sulfatase inhibitors may be useful asimmunosuppressive agents, and have been shown to enhance memory whendelivered to the brain.

Acne is a polyetiological disease caused by the interplay of numerousfactors, such as inheritance, sebum, hormones, and bacteria. The mostimportant causative factor in acne is sebum production; in almost allacne patients sebaceous glands are larger and more sebum is producedthan in persons with healthy skin. The development of the sebaceousgland and the extent of sebum production is controlled hormonally byandrogens; therefore, androgens play a crucial role in the pathogenesisof acne. In man, there are two major sources supplying androgens totarget tissues: (i) the gonades which secrete testosterone, (ii) theadrenals producing dehydroepiandrosterone (DHEA) which is secreted asthe sulfate conjugate (DHEAS). Testosterone and DHEAS are both convertedto the most active androgen, dihydrotestosterone (DHT), in the targettissue, e.g. in the skin. There is evidence that these pathways of localsynthesis of DHT in the skin are more important than direct supply withactive androgens from the circulation. Therefore, reduction ofendogeneous levels of androgens in the target tissue by specificinhibitors should be of therapeutic benefit in acne and seborrhoea.Furthermore, it opens the perspective to treat these disorders throughmodulation of local androgen levels by topical treatment, rather thaninfluencing circulating hormone levels by systemic therapies.

Androgenetic male alopecia is very common in the white races, accountingfor about 95% of all types of alopecia. Male-pattern baldness is causedby an increased number of hair follicles in the scalp entering thetelogen phase and by the telogen phase lasting longer. It is agenetically determined hair loss effected through androgens. Elevatedserum DHEA but normal testosterone levels have been reported in baldingmen compared with non-balding controls, implying that target tissueandrogen production is important in androgenetic alopecia.

Hirsutism is the pathological thickening and strengthening of the hairwhich is characterized by a masculine pattern of hair growth in childrenand women. Hirsutism is androgen induced, either by increased formationof androgens or by increased sensitivity of the hair follicle toandrogens. Therefore, a therapy resulting in reduction of endogeneouslevels of androgens and/or estrogens in the target tissue (skin) shouldbe effective in acne, androgenetic alopecia and hirsutism.

As described above, DHT, the most active androgen, is synthesized in theskin from the abundant systemic precursor DHEAS and the first step inthe metabolic pathway from DHEAS to DHT is desulfatation of DHEAS by theenzyme steroid sulfatase to produce DHEA. The presence of the enzyme inkeratinocytes and in skin-derived fibroblasts has been described. Thepotential use of steroid sulfatase inhibitors for the reduction ofendogenous levels of steroid hormones in the skin was confirmed usingknown steroid sulfatase inhibitors, such as estrone 3-O-sulfamate and4-methylumbelliferyl-7-O-sulfamate. We have found that inhibitors ofplacental steroid sulfatase also inhibit steroid sulfatase prepared fromeither a human keratinocyte (HaCaT) or a human skin-derived fibroblastcell line (1BR3GN). Such inhibitors were also shown to block steroidsulfatase in intact monolayers of the HaCaT keratinocytes.

Therefore, inhibitors of steroid sulfatase may be used to reduceandrogen and estrogen levels in the skin. They can be used as inhibitorsof the enzyme steroid sulfatase for the local treatment ofandrogen-dependent disorders of the pilosebaceous unit (such as acne,seborrhoea, androgenetic alopecia, hirsutism) and for the localtreatment of squamous cell carcinoma.

Furthermore non-steroidal steroid sulfatase inhibitors are expected tobe useful for the treatment of disorders mediated by the action ofsteroid hormones in which the steroidal products of the sulfatasecleavage play a role. Indications for these new kind of inhibitorsinclude androgen-dependent disorders of the pilosebaceous unit (such asacne, seborrhea, androgenetic alopecia, hirsutism); estrogen- orandrogen-dependent tumors, such as squamous cell carcinoma andneoplasms, e.g. of the breast, endometrium, and prostate; inflammatoryand autoimmune diseases, such as rheumatoid arthritis, type I and IIdiabetes, systemic lupus erythematosus, multiple sclerosis, myasteniagravis, thyroiditis, vasculitis, ulcerative colitis, and Crohn'sdisease, psoriasis, contact dermatitis, graft versus host disease,eczema, asthma and organ rejection following transplantation. Steroidsulfatase inhibitors are also useful for the treatment of cancer,especially for the treatment of estrogen- and androgen-dependentcancers, such as cancer of the breast and endometrium and squamous cellcarcinoma, and cancer of the prostata. Steroid sulfatase inhibitors arealso useful for the enhancement of cognitive function, especially in thetreatment of senile dementia, including Alzheimer's disease, byincreasing the DHEAS levels in the central nervous system.

Activities of compounds in inhibiting the activity of steroid sulfatasemay be shown in the following test systems:

Purification of Human Steroid Sulfatase

Human placenta is obtained freshly after delivery and stripped ofmembranes and connective tissues. For storage, the material is frozen at−70° C. After thawing, all further steps are carried out at 4° C., whilepH values are adjusted at 20° C. 400 g of the tissue is homogenized in1.2 l of buffer A (50 mM Tris-HCl, pH 7.4, 0.25 M sucrose). Thehomogenate obtained is centrifuged at 10,000×g for 45 minutes. Thesupernatant is set aside and the pellet obtained is re-homogenized in500 ml of buffer A. After centrifugation, the two supernatants obtainedare combined and subjected to ultracentrifugation (100,000×g, 1 hour).The pellet obtained is resuspended in buffer A and centrifugation isrepeated. The pellet obtained is suspended in 50 ml of 50 mM Tris-HCl,pH 7.4 and stored at −20° C. until further work-up.

After thawing, microsomes are collected by ultracentrifugation (asdescribed above) and are suspended in 50 ml of buffer B (10 mM Tris-HCl,pH 7.0, 1 mM EDTA, 2 mM 2-mercaptoethanol, 1% Triton X-100, 0.1%aprotnin). After 1 hour on ice with gentle agitation, the suspension iscentrifuged (100,000×g, 1 hour). The supernatant containing the enzymeactivity is collected and the pH is adjusted to 8.0 with 1 M Tris. Thesolution obtained is applied to a hydroxy apatite column (2.6×20 cm) andequilibrated with buffer B, pH 8.0. The column is washed with buffer Bat a flow rate of 2 ml/min. The activity is recovered in theflow-through. The pool is adjusted to pH 7.4 and subjected tochromatography on a concanavalin A sepharose column (1.6×10 cm)equilibrated in buffer C (20 mM Tris-HCl, pH 7:4, 0.1% Triton X-100, 0.5M NaCl). The column is washed with buffer C, and the bound protein iseluted with 10% methyl mannoside in buffer C. Active fractions arepooled and dialysed against buffer D (20 mM Tris-HCl, pH 8.0, 1 mM EDTA,0.1% Triton X-100, 10% glycerol (v/v)).

The retentate obtained is applied to a blue sepharose column (0.8×10 cm)equilibrated with buffer D; which column is washed and elution iscarried out with a linear gradient of buffer D to 2 M NaCl in buffer D.Active fractions are pooled, concentrated as required (Centricon 10),dialysed against buffer D and stored in aliquots at −20° C.

Assay of Human Steroid Sulfatase

It is known that purified human steroid sulfatase not only is capable tocleave steroid sulfates, but also readily cleaves aryl sulfates such as4-methylumbelliferyl sulfate which is used in the present test system asan activity indicator. Assay mixtures are prepared by consecutivelydispensing the following solutions into the wells of white microliterplates:

-   1) 50 μl substrate solution (1.5 mM 4-methylumbelliferyl sulfate in    0.1 M Tris-HCl, pH 7.5)-   2) 50 μl test compound dilution in 0.1 M Tris-HCl, pH 7.5, 0.1%    Triton X-100 (stock solutions of the test compounds are prepared in    DMSO; final concentrations of the solvent in the assay mixture not    exceeding 1%)-   3) 50 μl enzyme dilution (approximately 12 enzyme units/ml)

We define one enzyme unit as the amount of steroid sulfatase thathydrolyses 1 nmol of 4-methylumbelliferyl sulfate per hour at an initialsubstrate concentration of 500 μM in 0.1 M Tris-HCl, pH 7.5, 0.1% TritonX-100, at 37° C.

Plates are incubated at 37° C. for 1 hour. Then the reaction Is stoppedby addition of 100 μl 0.2 M NaOH. Fluorescence intensity is determinedin a Titertek Fluoroskan. II instrument with λ_(e)=355 nm and λ_(em)=460nm.

Calculation of Relative IC₅₀ Values

From the fluorescence intensity data (I) obtained at differentconcentrations (c) of the test compound in the human steroid sulfataseassay as described above, the concentration inhibiting the enzymaticactivity by 50% (IC₅₀) is calculated using the equation:$I = \frac{I_{100}}{1 + \left( {c/{IC}_{50}} \right)^{s}}$wherein I₁₀₀ is the intensity observed in the absence of inhibitor and sis a slope factor. Estrone sulfamate is used as a reference compound andits IC₅₀ value is determined in parallel to all other test compounds.Relative IC₅₀ values are defined as follows:${{rel}\quad{IC}_{50}} = \frac{{IC}_{50}\quad{of}\quad{test}\quad{compound}}{{IC}_{50}\quad{of}\quad{estrone}\quad{sulfamate}}$

According to our testing and calculation estrone sulfamate shows an IC₅₀value of approximately 60 nM.

The compounds of the present invention show activity in that describedassay (rel IC₅₀ in the range of 0.0046 to 350).

CHO/STS Assay

CHO cells stably transfected with human steroid sulfatase (CHO/STS) areseeded into microtiter plates. After reaching approximately 90%confluency, they are incubated overnight with graded concentrations oftest substances (e.g. compounds of the present invention). They are thenfixed with 4% paraformaldehyde for 10 minutes at room temperature andwashed 4 times with PBS, before incubation with 100 μl/well 0.5 mM4-methylumbelliferyl sulfate (MUS), dissolved in 0.1 M Tris-HCl, pH 7.5.The enzyme reaction is carried out at 37° C. for 30 minutes. Then 50μl/well stop solution (1 M Tris-HCl, pH 10.4) are added. The enzymereaction solutions are transferred to white plates (Microfluor, Dynex,Chantilly, Va.) and read in a Fluoroskan II fluorescence microtiterplate reader. Reagent blanks are subtracted from all values. For drugtesting, the fluorescence units (FU) are divided by the optical densityreadings after staining cellular protein with sulforhodamine B (OD₅₅₀),in order to correct for variations in cell number. IC₅₀ values aredetermined by linear interpolation between two bracketing points. Ineach assay with inhibitors, estrone 3-O-sulfamate is run as a referencecompound, and the IC₅₀ values are normalized to estrone 3-O-sulfamate(relative IC₅₀=IC₅₀ compound/IC₅₀ estrone 3-O-sulfamate).

The compounds of the present invention show activity in that describedassay (rel lC₅₀ in the range of 0.05 to 226).

Assay Using Human Skin Homogenate

Frozen specimens of human cadaver skin (about 100 mg per sample) areminced Into small pieces (about 1×1 mm) using sharp scissors. The piecesobtained are suspended in ten volumes (w/w) of buffer (20 mM Tris-HCl,pH 7.5), containing 0.1% Triton X-100. Test compounds (e.g. compounds ofthe present invention) are added at graded concentrations from stocksolutions In ethanol or DMSO. Second, DHEAS as the substrate is added (1μC/ml [3H]DHEAS, specific activity: about 60 Ci/mmol, and 20 μMunlabeled DHEAS).

Samples are incubated for 18 hrs at 37° C. At the end of the Incubationperiod, 50 μl of 1 M Tris, pH 10.4 and 3 ml of toluene are added. A 1-mlaliquot of the organic phase is removed and subjected to liquidscintillation counting. The determined dpm-values in the aliquots areconverted to nmol of DHEA cleaved per g of skin per hour.

The compounds of the present invention show activity in that describedassay (IC₅₀ in the range of 0.03 to 10 μM).

A preferred compound of the present invention includes a compound ofExample 1. That compound show in the Human Steroid Sulfatase Assay a relIC₅₀ in the range of 0.0046 to 0.071, in the CHO/STS Assay a rel IC₅₀ inthe range of 0.02 to 0.39, and in the Assay Using Human Skin Homogenateof an IC₅₀ in the range of 0.03 to 0.27 μM and is thus a highly activesteroide sulfatase inhibitor.

The compounds of the present invention show activity in test systems asdefined above. A compound of the present invention in salt and/orsolvate form exhibits the same order of activity as a compound of thepresent invention in free and/or non-solvated form.

The compounds of the present invention are therefore indicated for useas steroid sulfatase inhibitors in the treatment of disorders mediatedby the action of steroid sulfatase, e.g. including androgen-dependentdisorders of the pilosebaceous unit, such as

-   -   acne,    -   seborrhea,    -   androgenetic alopecia,    -   hirsutism;    -   cancers, such as estrogen and androgen-dependent cancers;    -   cognitive dysfunctions, such as senile dementia including        Alzheimer's disease.

The compounds of the present invention are preferably used in thetreatment of acne, seborrhea, androgenetic alopecia, hirsutism;estrogen, e.g. and androgen-dependent cancers, more preferably in thetreatment of acne. Treatment includes therapeutical treatment andprophylaxis.

In another aspect the present invention provides the use of a compoundof the present invention for the manufacture of a medicament, e.g. apharmaceutical composition, for the treatment of disorders mediated bythe action of steroid sulfatase, e.g. a disorder responsive to theinhibition of the action of steroid sulfatase, most preferably acne.

For such use a compound of the present invention includes one or morecompounds of the present invention, e.g. a combination of two or morecompounds of the present invention, preferably one.

In another aspect the present invention provides a compound of thepresent invention for use as a pharmaceutical.

In another aspect the present invention provides a method of treatmentof disorders mediated by the action of steroid sulfatase, such as acne,seborrhea, androgenetic alopecia, hirsutism; cancers, such as estrogenand androgen-dependent cancers; cognitive dysfunctions, such as seniledementia including Alzheimer's disease, preferably acne, which treatmentcomprises administering to a subject in need of such treatment aneffective amount of a compound of the present invention; e.g. in theform of a pharmaceutical composition.

Treatment includes treatment and prophylaxis.

For such treatment, the appropriate dosage will, of course, varydepending upon, for example, the chemical nature and the pharmakokineticdata of a compound of the present invention employed, the individualhost, the mode of administration and the nature and severity of theconditions being treated. However, in general, satisfactory results maybe obtained if the compounds are administered at a daily dose of fromabout 0.1 mg/kg to about 100 mg/kg animal body weight, e.g. convenientlyadministered In divided doses two to four times daily. For most mammalsthe total daily dosage is from about 5 mg to about 5000 mg, convenientlyadministered, for example, in divided doses up to four times a day or inretarded form. Unit dosage forms comprise, e.g. from about 1.25 mg toabout 2000 mg of a compound of a present invention in admixture with atleast one pharmaceutically acceptable excipient, e.g. carrier, diluent.

A compound of the present invention may be administered by anyconventional route, for example enterally, e.g. Including nasal, buccal,rectal, oral, administration; parenterally, e.g. including intravenous,intramuscular, subcutanous administration; or topically; e.g. includingepicutaneous, intranasal, intratracheal administration; e.g. in form ofcoated or uncoated tablets, capsules, injectable solutions orsuspensions, e.g. in the form of ampoules, vials, in the form ofointments, creams, gels, pastes, inhaler powder, foams, tinctures, lipsticks, drops, sprays, e.g. or in the form of suppositories. Thecompounds of the present invention may be administered in the form of apharmaceutically acceptable salt, or in free form; optionally in theform of a solvate. The compounds of the present invention in the form ofa salt exhibit the same order of activity as the compounds of thepresent invention in free form; optionally in the form of a solvate.

A compound of the present invention may be used for pharmaceuticaltreatment according to the present invention alone, or in combinationwith one or more other pharmaceutically active agents. Such otherpharmaceutically active agents include e.g. retinoids, e.g. retinoicacid, such as isotretinoin; tretinoin (Roche); adapalene(6-[3-(1-adamantyl)-4-methoxyphenyl]-2-naphthoic acid); oralcontraceptives, e.g. 19-nor-17a-pregna-1,3,5(10)-trien-20-in-3,17-diol,6-Chlor-17-hydroxy-1a,2a-methylen-4,6-pregnadien-3,20-dion, such asDiane® (Schering), antibacterials, such as erythromycins, includingerythromycin A, azithromycin, clarithromycin, roxythromycin;tetracyclines, lincosamid-antibiotics, such as clindamycin (methyl7-chlor-6,7,8-tridesoxy-6-(trans-1-methylpropyl-L-2-pyrrolidin-carboxamido)-1-thio-L-threo-a-D-galacto-octopyranosid),azelaic acid (nonanedionic acid), nadifloxacin; dapsone, benzoylperoxide; keratolytics, such as salicylic acid; anti-inflammatoryagents, such as corticosteroids, pimecrolimus; steroid 5α-reductaseinhibitors. For the treatment of breast and endometrial cancer furtherpharmaceutically active agents include aromatase inhibitors, such asanastrozole, letrozole, exemestane.

Combinations include

-   -   fixed combinations, in which two or more pharmaceutically active        agents are in the same pharmaceutical composition,    -   kits, in which two or more pharmaceutically active agents in        separate compositions are sold in the same package, e.g. with        instruction for co-administration; and    -   free combinations in which the pharmaceutically active agents        are packaged separately, but instruction for simultaneous or        sequential administration are given.

In another aspect the present invention provides a pharmaceuticalcomposition comprising a compound of the present invention inassociation with at least one pharmaceutical excipient, e.g. appropriatecarrier and/or diluent, e.g. including fillers, binders, disintegrators,flow conditioners, lubricants, sugars and sweeteners, fragrances,preservatives, stabilizers, wetting agents and/or emulsifiers,solubilizers, salts for regulating osmotic pressure, buffers.

In another aspect the present invention provides a pharmaceuticalcomposition according to the present invention, further comprisinganother pharmaceutically active agent.

Such pharmaceutical compositions may be manufactured according, e.g.analogously to a method as conventional, e.g. by mixing, granulating,coating, dissolving or lyophilizing processes.

In the following example references to temperature are in degreeCentigrade and are uncorrected. ¹HNMR are carried out in CDCl₃ if nototherwise indicated.

The following abbreviations are used:

-   m.p.: melting point-   EtAc: ethyl acetate-   BOC: tert.butyloxycarbonyl-   DMF: N,N-dimethylformamide-   RT: room temperature-   DMSO: dimethylsulfoxide

EXAMPLE 1N-[[4-[1-(2-Aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-amino]-sulfonyl-3,5-bis-trifluoromethyl-benzamideA) 4-[N-(Aminosulfonyl)benzylamino]-piperidine-1-carboxylic acidtert.butylester

A solution of 2.05 g of 4-benzylamino-piperidine-1-carboxylic acidtert.-butyl ester and 3.39 g of sulfamide in 100 ml of1,2-dimethoxy-ethane is refluxed for ca. 10 hours. From the mixtureobtained solvent is evaporated, the evaporation residue obtained istreated with EtAc and unsoluble parts are filtered off. The filtrateobtained is subjected to chromatography on silica gel.4-[N-(Aminosulfonyl)-benzylamino]-piperidine-1-carboxylic acidtert.butylester is obtained. mp 140-142°. ¹H-NMR (DMSO) δ: 7.30-7.41(s/m, 9+2H), 1.64 (m, 2H), 2.53-2.72 (broad, 2H), 3.71 (tt, 1H),8.83-8.96 (broad, 2H), 4.26 (s, 2H, CH2), 6.86 (bs, 2H, NH), 7.19 (t,1H), 7.28 (t, 1H), 7.36 (d, 1H). ¹³C-NMR (DMSO) δ: 28.51, 30.13, 43.35,47.11, 56.52, 79.12, 127.06, 127.48, 128.49, 140.59, 154.156.

B)N-(1-BOC-4-Piperidinyl-benzylamino)-sulfonyl-3,5-bis-trifluoromethyl-benzamide

280 mg of diisopropyl-ethylamine and 1.28 ml of n-propylphosphonicanhydride (50% solution in DMF) are added to a solution of 400 mg of4-[N-(aminosulfonyl)-benzylamino]-piperidine-1-carboxylic acidtert.butylester and 560 mg of 3,5-bis-(trifluoromethyl)benzoic acid in15 ml of DMF. The mixture obtained is stirred for ca. 4 days at RT,solvent is evaporated and the evaporation residue is treated with EtAcand washed with 1 N HCl, sat. NaHCO₃ solution and brine and dried. Fromthe dried residue solvent is evaporated and the evaporation residueobtained is subjected to chromatograpy on silica gel.N-(1-BOC-4-Piperidinyl-benzylamino)-sulfonyl-3,5-bis-4-trifluoromethyl-benzamideis obtained.

¹H-NMR (DMSO) δ: 1.28-1.40 (m, 9+2H), 1.60 (m, 2H), 2.54-2.70 (m, 2H),3.80-3.94 (m, 2+1H), 4.55 (s, 2H,CH2), 7.18 (t, 1H), 7.27 (d, 1H), 7.37(d, 1H), 8.26 (s, 1H), 8.46 (s, 2H), 12.6 (broad, 1H, NH).

C)N-(4-Piperidinyl-benzylamino)sulfonyl-3,5-bis-trifluoromethyl-benzamidein the form of a hydrochloride

A solution of 650 mg ofN-(1-BOC-4-piperidinyl-benzylamino)-sulfonyl-3,5-bis-trifluoromethyl-benzamidein 5 ml of CH₂Cl₂ is treated with 60 ml of 3 N HCl (gas) indiethylether. The mixture obtained is stirred for ca. 12 hours andsolvent is evaporated.N-(4-Piperidinyl-benzylamino)-sulfonyl-3,5-bis-trifluoromethyl-benzamidein the form of a hydrochloride is obtained. ¹H-NMR (DMSO) δ: 1.72-1.86(m, 4H), 2.90 (m, 2H), 3.20-3.3 (m, 2H), 4.15 (m, 1H), 4.60 (s, 2H,CH2), 7.31 (t, 1H), 7.40 (t, 1H), 7.48 (d, 1H), 8.38 (s, 1H), 8.48 (s,2H), 8.48 and 8.75 (broad, 2H, NH).

D)N-[[4-[1-(2-Aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-benzylamino]-sulfonyl-3,5-bis-trifluoromethyl-benzamide

430 mg of 2-fluoro-4-trifluoromethyl-benzamide and 430 mg of K₂CO₃ areadded to a solution of 570 mg ofN-(4-piperidinyl-benzylamino)-sulfonyl-3,5-bis-trifluoromethyl-benzamidehydrochloride in 15 ml of DMSO. The mixture obtained is stirred for ca.4 hours at 150° and K₂CO₃ is removed by filtration. Solvent from thefiltrate obtained is evaporated and the evaporation residue is subjectedto chromatography on silica gel.N-[[4-[1-(2-Aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-benzylamino]-sulfonyl-3,5-bis-trifluoromethyl-benzamideis obtained. ¹H-NMR (DMSO) δ: 1.60-1.76 (m, 4H), 2.72 (m, 2H), 3.13 (m,2H), 3.78 (m, 1H), 4.55 (s, 2H, CH2), 7.14 (t, 1H), 7.23 (t, 1H), 7.26(d, 1H), 7.32 (d, 1H), 7.41 (d, 1H), 7.59 (broad, 1H, NH), 7.68 (d, 1H),8.08 (broad, 1H, NH), 8.12 (s, 1H), 8.45 (s, 2H).

E)N-[[4-[1-(2-Aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-amino]-sulfonyl-3,5-bis-trifluoromethyl-benzamide

830 mg ofN-[[4-[1-(2-aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-benzylamino]-sulfonyl-3,5-bis-trifluoromethyl-benzamideare dissolved in 20 ml of MeOH and the mixture obtained is hydrogenatedfor ca. 12 hours at 40° in the presence of 10% palladium on charcoal.The catalyst is filtered off and the filtrate obtained is subjected tochromatography on silica gel.N-[[4-[1-(2-Aminocarbonyl-5-trifluoromethyl-phenyl)]-piperidinyl]-amino]-sulfonyl-3,5-bis-trifluoromethyl-benzamideis obtained. mp 192-195° C. ¹H-NMR (DMSO) δ: 1.58 (m, 2H), 1.94 (m, 2H),2.77 (m, 2H), 3.11-3.21 (m, 3H), 5.8 (broad, 1H, NH), 7.32 (s, 1H), 7.36(d, 1H), 7.69 (broad, 1H), 7.74 (d, 1H), 8.16 (s, 1H), 8.22 (broad, 1H),8.48 (s, 2H).

Analogously to the method as described in example 1 but usingappropriate starting materials compounds of formula

are obtained, wherein R_(3ss) and R_(4ss) are as defined in TABLE 1having a melting point as defined in TABLE 1: TABLE 1 Example R_(3ss)R_(4ss) m.p. 1

192-195° 2

215-220° 3

149-152° 4

190°decom- position

EXAMPLE 5 3,5-Bis(trifluoromethyl)benzoyl-sulfamic acidN-BOC-piperazineamide A) N-BOC-piperazine-N′-sulfamate

612 mg of amidosulfonyl chloride in 20 ml of CH₂Cl₂ are added to asolution of 1 g of BOC-piperazine and 1.085 g of triethylamine. Themixture obtained is stirred for ca. 6 hours at 0° and allowed to warm toRT overnight. Solvent of the mixture obtained is evaporated, theevaporation residue obtained is dissolved in ethyl acetate, washed withH₂O, dried and concentrated. The concentration residue obtained issubjected to chromatography on silicagel. N-BOC-piperazine-N′-sulfamatein crystalline form is obtained. m.p.: 167-171°.

B) 3,5-Bis(trifluoromethyl)benzoyl-sulfamic acid N-BOC-piperazineamide

To a solution of 195 mg of N-BOC-piperazine-N′-sulfamate and 148 mg oftriethylamine are added 406 mg of3,5-bis(trifluoromethyl)benzoylchloride in 50 ml of CH₂Cl₂. The mixtureobtained is stirred for ca. 20 hours at RT, solvent is evaporated, theevaporation residue obtained is dissolved in EtAc, washed with H₂O,dried and concentrated. The concentration residue obtained is subjectedto chromatograpy on silica gel. 3,5-bis(trifluoromethyl)benzoyl-sulfamicacid N-BOC-piperazineamide in crystalline form are obtained. m.p.:185-190°. ¹H-NMR □: 1.4 (s, 9H. BOC), 3.0-3.2 (m, 4H), 3.3-3.5 (m, 4H),7.78 (s, 1H), 8.28 (s, 2H).

Analogously to the method as described In example 5 but usingappropriate starting materials compounds of formula

wherein R_(3s) and R_(4s) are defined in TABLE 2, having the meltingpoint as set out in TABLE 2 are obtained: TABLE 2 Example R_(3s) R_(4s)m.p. 5

185-190° 6

172-175° 7

200-203° 8

208-211° 9

63-6720 10

205-208° 11

195-197° 12

193-197° 13

154-157° 14

186-189°

1. A compound of formula

wherein either R₁ and R₂ together with the nitrogen atom to which theyare attached are piperazinyl, wherein the second nitrogen atom issubstituted by (C₁₋₆)alkoxycarbonyl or by (C₆₋₁₈)aryl, which (C₆₋₁₈)arylis substituted by one or more halogen, (C₁₋₆)haloalkyl, e.g. CF₃,aminocarbonyl, or R₁ is hydrogen and R₂ is piperidinyl, attached via acarbon atom of the piperidinyl ring, wherein the nitrogen atom issubstituted by (C₁₋₆)alkoxycarbonyl or by (C₆₋₁₈)aryl, and R₃ is(C₆₋₁₈)aryl or (C₆₋₁₈)aryl(C₁₋₄)alkyl, which aryl is substituted by oneor more halogen, aminocarbonyl, or (C₁₋₆)haloalkyl.
 2. A compound ofclaim 1 selected from the group consisting of compounds of formula

wherein a. R_(3ss) is 3,5-bis(trifluoromethyl)phenyl and R_(4ss) is2-aminocarbonyl-5-trifluoromethylphenyl, b. R_(3ss) is2,3-dichlorophenyl, R_(4ss) is 2-aminocarbonyl-5-trifluoromethylphenyl,c. R_(3ss) is 3,5-dichlorophenyl R_(4ss) is2-aminocarbonyl-5-trifluoromethylphenyl, and d. R_(3ss) is3,5-bis(trifluoromethyl)phenyl and R_(4ss) is tert.butoxycarbonyl.
 3. Acompound of claim 1 which is a compound of formula


4. A compound of claim 1 selected from the group consisting of compoundsof formula

wherein a. R_(3s) is 3,5-bis(trifluoromethyl)phenyl and R_(4s) istert.butoxycarbonyl, b. R_(3s) is 2,3-dichlorophenyl and R_(4s) istert.butoxycarbonyl, c. R_(3s) is 3,5-dichlorophenyl and R_(4s) istert.butoxycarbonyl, d. R_(3s) is 3,5-bis(trifluoromethyl)phenyl andR_(4s) is benzyloxycarbonyl, e. R_(3s) is 2,3-dichlorophenyl and R_(4s)is benzyloxycarbonyl, f. R_(3s) is 3,5-dichlorophenyl and R_(4s) isbenzyloxycarbonyl, g. R_(3s) is 3,5-dichlorophenyl and R_(4s) isbenzyloxycarbonyl, h. R_(3s) is 3,5-bis(trifluoromethyl)phenyl andR_(4s) is 2-aminocarbonyl-5-trifluoromethylphenyl, i. R_(3s) is3,5-dichlorophenyl and R_(4s) is is2-aminocarbonyl-5-trifluoromethylphenyl, j. R_(3s) is 2,3-dichlorophenyland R_(4s) is is 2-aminocarbonyl-5-trifluoromethylphenyl, and k. R_(3s)is 2-(3,5-bis(trifluoromethyl)phenyl)ethyl and R_(4s) is istert.butoxycarbonyl.
 5. A compound of claim 1 in the form of a salt. 6.Use of a compound of claim 1 for the manufacture of a medicament for thetreatment of disorders mediated by the action of steroid sulfatase.
 7. Acompound of claim 1 for use as a pharmaceutical.
 8. A method oftreatment of disorders mediated by the action of steroid sulfatase whichtreatment comprises administering to a subject in need of such treatmentan effective amount of a compound of claim
 1. 9. A pharmaceuticalcomposition comprising a compound of claim 1 in association with atleast one pharmaceutical excipient.
 10. A pharmaceutical compositionaccording to claim 9, further comprising another pharmaceutically activeagent.